Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

The Ndc80/HEC1 complex is a contact point for kinetochore-microtubule attachment

Nature Structural & Molecular Biology volume 14, pages 54–59 (2007)Cite this article

Abstract

Kinetochores are multicomponent assemblies that connect chromosomal centromeres to mitotic-spindle microtubules. The Ndc80 complex is an essential core element of kinetochores, conserved from yeast to humans. It is a rod-like assembly of four proteins— Ndc80p (HEC1 in humans), Nuf2p, Spc24p and Spc25p. We describe here the crystal structure of the most conserved region of HEC1, which lies at one end of the rod and near the N terminus of the polypeptide chain. It folds into a calponin-homology domain, resembling the microtubule-binding domain of the plus-end-associated protein EB1. We show that an Ndc80p-Nuf2p heterodimer binds microtubules in vitro. The less conserved, N-terminal segment of Ndc80p contributes to the interaction and may be a crucial regulatory element. We propose that the Ndc80 complex forms a direct link between kinetochore core components and spindle microtubules.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Structure of the HEC1 globular domain, compared with the EB1 microtubule-binding domain.
Figure 2: Surface properties of HEC1_CH.
Figure 3: Microtubule binding by 2NG and 2NGΔN.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank

References

  1. Koshland, D.E., Mitchison, T.J. & Kirschner, M.W. Polewards chromosome movement driven by microtubule depolymerization in vitro. Nature 331, 499–504 (1988).

    Article  CAS  PubMed  Google Scholar 

  2. Cleveland, D.W., Mao, Y. & Sullivan, K.F. Centromeres and kinetochores: from epigenetics to mitotic checkpoint signaling. Cell 112, 407–421 (2003).

    Article  CAS  PubMed  Google Scholar 

  3. McAinsh, A.D., Tyell, J.D. & Sorger, P.K. Structure, function, and regulation of budding yeast kinetochores. Annu. Rev. Cell Dev. Biol. 19, 519–539 (2003).

    Article  CAS  PubMed  Google Scholar 

  4. Joglekar, A.P., Bouck, D.C., Molk, J.N., Bloom, K.S. & Salmon, E.D. Molecular architecture of a kinetochore-microtubule attachment site. Nat. Cell Biol. 8, 581–585 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Roos, U.P. Light and electron microscopy of rat kangaroo cells in mitosis. II. Kinetochore structure and function. Chromosoma 41, 195–220 (1973).

    Article  CAS  PubMed  Google Scholar 

  6. Chikashige, Y. et al. Composite motifs and repeat symmetry in S. pombe centromeres: direct analysis by integration of NotI restriction sites. Cell 57, 739–751 (1989).

    Article  CAS  PubMed  Google Scholar 

  7. Clarke, L. & Carbon, J. Isolation of the centromere-linked CDC10 gene by complementation in yeast. Proc. Natl. Acad. Sci. USA 77, 2173–2177 (1980).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Cottarel, G., Shero, J.H., Hieter, P. & Hegemann, J.H. A 125-base-pair CEN6 DNA fragment is sufficient for complete meiotic and mitotic centromere functions in Saccharomyces cerevisiae. Mol. Cell. Biol. 9, 3342–3349 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Peterson, J.B. & Ris, H. Electron-microscopic study of the spindle and chromosome movement in the yeast Saccharomyces cerevisiae. J. Cell Sci. 22, 219–242 (1976).

    CAS  PubMed  Google Scholar 

  10. McDonald, K.L., O'Toole, E.T., Mastronarde, D.N. & McIntosh, J.R. Kinetochore microtubules in PTK cells. J. Cell Biol. 118, 369–383 (1992).

    Article  CAS  PubMed  Google Scholar 

  11. Winey, M. et al. Three-dimensional ultrastructural analysis of the Saccharomyces cerevisiae mitotic spindle. J. Cell Biol. 129, 1601–1615 (1995).

    Article  CAS  PubMed  Google Scholar 

  12. Meraldi, P., McAinsh, A.D., Rheinbay, E. & Sorger, P.K. Phylogenetic and structural analysis of centromeric DNA and kinetochore proteins. Genome Biol. 7, R23 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  13. Kline-Smith, S.L., Sandall, S. & Desai, A. Kinetochore-spindle microtubule interactions during mitosis. Curr. Opin. Cell Biol. 17, 35–46 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Deluca, J.G. et al. Hec1 and nuf2 are core components of the kinetochore outer plate essential for organizing microtubule attachment sites. Mol. Biol. Cell 16, 519–531 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. He, X., Rines, D.R., Espelin, C.W. & Sorger, P.K. Molecular analysis of kinetochore-microtubule attachment in budding yeast. Cell 106, 195–206 (2001).

    Article  CAS  PubMed  Google Scholar 

  16. Janke, C., Ortiz, J., Tanaka, T.U., Lechner, J. & Schiebel, E. Four new subunits of the Dam1-Duo1 complex reveal novel functions in sister kinetochore biorientation. EMBO J. 21, 181–193 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. McCleland, M.L. et al. The highly conserved Ndc80 complex is required for kinetochore assembly, chromosome congression, and spindle checkpoint activity. Genes Dev. 17, 101–114 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Li, Y. et al. The mitotic spindle is required for loading of the DASH complex onto the kinetochore. Genes Dev. 16, 183–197 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Tirnauer, J.S., Grego, S., Salmon, E.D. & Mitchison, T.J. EB1-microtubule interactions in Xenopus egg extracts: role of EB1 in microtubule stabilization and mechanisms of rargeting to microtubules. Mol. Biol. Cell 13, 3614–3626 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Wei, R.R., Sorger, P.K. & Harrison, S.C. Molecular organization of the Ndc80 complex, an essential kinetochore component. Proc. Natl. Acad. Sci. USA 102, 5363–5367 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Gillett, E.S., Espelin, C.W. & Sorger, P.K. Spindle checkpoint proteins and chromosome-microtubule attachment in budding yeast. J. Cell Biol. 164, 535–546 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Ciferri, C. et al. Architecture of the human ndc80-hec1 complex, a critical constituent of the outer kinetochore. J. Biol. Chem. 280, 29088–29095 (2005).

    Article  CAS  PubMed  Google Scholar 

  23. Wei, R.R. et al. Structure of a central component of the yeast dinetochore: rhe Spc24p/Spc25p globular domain. Structure 14, 1003–1009 (2006).

    Article  CAS  PubMed  Google Scholar 

  24. Hayashi, I. & Ikura, M. Crystal structure of the amino-terminal microtubule-binding domain of end-binding protein 1 (EB1). J. Biol. Chem. 278, 36430–36434 (2003).

    Article  CAS  PubMed  Google Scholar 

  25. Holm, L. & Sander, C. Protein structure comparison by alignment of distance matrices. J. Mol. Biol. 233, 123–138 (1993).

    Article  CAS  PubMed  Google Scholar 

  26. Goldsmith, S.C. et al. The structure of an actin-crosslinking domain from human fimbrin. Nat. Struct. Biol. 4, 708–712 (1997).

    Article  CAS  PubMed  Google Scholar 

  27. Gimona, M., Djinovic-Carugo, K., Kranewitter, W.J. & Winder, S.J. Functional plasticity of CH domains. FEBS Lett. 513, 98–106 (2002).

    Article  CAS  PubMed  Google Scholar 

  28. Matsudaira, P. Modular organization of actin crosslinking proteins. Trends Biochem. Sci. 16, 87–92 (1991).

    Article  CAS  PubMed  Google Scholar 

  29. Klein, M.G. et al. Structure of the actin crosslinking core of fimbrin. Structure 12, 999–1013 (2004).

    Article  CAS  PubMed  Google Scholar 

  30. Borrego-Diaz, E. et al. Crystal structure of the actin-binding domain of [alpha]-actinin 1: Evaluating two competing actin-binding models. J. Struct. Biol. 155, 230–238 (2006).

    Article  CAS  PubMed  Google Scholar 

  31. Tirnauer, J.S. & Bierer, B.E. EB1 proteins regulate microtubule dynamics, cell polarity, and chromosome stability. J. Cell Biol. 149, 761–766 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Slep, K.C. et al. Structural determinants for EB1-mediated recruitment of APC and spectraplakins to the microtubule plus end. J. Cell Biol. 168, 587–598 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Tanaka, T.U. et al. Evidence that the Ipl1-Sli15 (Aurora kinase-INCENP) complex promotes chromosome bi-orientation by atering kinetochore-spindle pole connections. Cell 108, 317–329 (2002).

    Article  CAS  PubMed  Google Scholar 

  34. Lampson, M.A., Renduchitala, K., Khodjakov, A. & Kapoor, T.M. Correcting improper chromosome-spindle attachments during cell division. Nat. Cell Biol. 6, 232–237 (2004).

    Article  CAS  PubMed  Google Scholar 

  35. Cheeseman, I.M. et al. Phospho-regulation of kinetochore-microtubule attachments by the Aurora kinase Ipl1p. Cell 111, 163–172 (2002).

    Article  CAS  PubMed  Google Scholar 

  36. Nousiainen, M., Sillje, H.H.W., Sauer, G., Nigg, E.A. & Korner, R. Phosphoproteome analysis of the human mitotic spindle. Proc. Natl. Acad. Sci. USA 103, 5391–5396 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. DeLuca, J.G. et al. Kinetochore microtubule dynamics and attachment stability are regulated by Hec1. Cell 127, 969–982 (2006).

    Article  CAS  PubMed  Google Scholar 

  38. Cheeseman, I.M. et al. The conserved KMN network constitutes the core microtubule- binding site of the kinetochore. Cell 127, 987–997 (2006).

    Article  Google Scholar 

  39. Tanaka, K. et al. Molecular mechanisms of kinetochore capture by spindle microtubules. Nature 434, 987–994 (2005).

    Article  CAS  PubMed  Google Scholar 

  40. Miranda, J.J., De Wulf, P., Sorger, P. & Harrison, S.C. The yeast DASH complex forms closed rings on microtubules. Nat. Struct. Mol. Biol. 12, 138–143 (2005).

    Article  CAS  PubMed  Google Scholar 

  41. Westermann, S. et al. Formation of a dynamic kinetochore-microtubule interface through assembly of the Dam1 ring complex. Mol. Cell 17, 277–290 (2005).

    Article  CAS  PubMed  Google Scholar 

  42. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  PubMed  Google Scholar 

  43. Terwilliger, T.C. & Berendzen, J. Automated MAD and MIR structure solution. Acta Crystallograph. D Biol. Crystallogr. 55, 849–861 (1999).

    Article  CAS  Google Scholar 

  44. Terwilliger, T. Maximum-likelihood density modification. Acta Crystallograph. D Biol. Crystallogr. 56, 965–972 (2000).

    Article  CAS  Google Scholar 

  45. Jones, T.A., Zou, J.Y., Cowan, S.W. & Kjeldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  PubMed  Google Scholar 

  46. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  PubMed  Google Scholar 

  47. Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).

    Article  CAS  PubMed  Google Scholar 

  48. Roger, B., Al-Bassam, J., Dehmelt, L., Milligan, R.A. & Halpain, S. MAP2c, but not Tau, binds and bundles F-actin via its microtubule binding domain. Curr. Biol. 14, 363–371 (2004).

    Article  CAS  PubMed  Google Scholar 

  49. Honig, B. & Nicholls, A. Classical electrostatics in biology and chemistry. Science 268, 1144–1149 (1995).

    Article  CAS  PubMed  Google Scholar 

  50. Chenna, R. et al. Multiple sequence alignment with the Clustal series of programs. Nucleic Acids Res. 31, 3497–3500 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors thank J.J. Miranda for help with the microtubule cosedimentation assay, D. Panne for assistance with X-ray data collection and E. Salmon, J. Deluca, N. Larsen, P.R. Ohi and V. Draviam for critique of the manuscript. We acknowledge the Advanced Light Source at Lawrence Berkeley National Laboratory. J.A.-B. is a recipient of the American Cancer Society postdoctoral fellowship. S.C.H. is an investigator of the Howard Hughes Medical Institute.

Author information

Authors and Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 250 Longwood Avenue, Boston, 02115, Massachusetts, USA

    Ronnie R Wei & Jawdat Al-Bassam

  2. Howard Hughes Medical Institute, Harvard Medical School, 250 Longwood Avenue, Boston, 02115, Massachusetts, USA

    Ronnie R Wei & Stephen C Harrison

Authors
  1. Ronnie R Wei
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jawdat Al-Bassam
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Stephen C Harrison
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

R.R.W. contributed to the design and execution of the structure determination of HEC1_CH, the microtubule cosedimentation experiments and manuscript preparation. J.A.-B. and R.R.W. contributed the negative-stain electron microscopy. S.C.H. guided the project and helped prepare the manuscript.

Corresponding author

Correspondence to Stephen C Harrison.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Comparison of relative microtubule-binding affinity of Ndc80 and EB1. (PDF 68 kb)

Supplementary Fig. 2

Ndc80p N-terminal segment alone does not bind to microtubules. (PDF 11 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wei, R., Al-Bassam, J. & Harrison, S. The Ndc80/HEC1 complex is a contact point for kinetochore-microtubule attachment. Nat Struct Mol Biol 14, 54–59 (2007). https://doi.org/10.1038/nsmb1186

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb1186

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing